Project 4 focuses on the ability of circadian genes in the ventral tegmental area (VT A)-nucleus accumbens (NAc) pathway to regulate mood and motivational state. This is related to the knowledge that abnormal mood and other symptoms in many patients with depression show prominent circadian oscillations. We have demonstrated that NPAS2 (neuronal PAS domain protein 2), a transcription factor highly homologous to Clock, regulates an animal's responsiveness to emotional stimuli, including their activity in animal models of depression. Interestingly, NPAS2 is not expressed in the suprachiasmatic nucleus (SCN), a hypothalamic region important for circadian oscillations and their entrainment by environmental lighting. Rather, the highest expression of NPAS2 is seen in the NAc. Our hypothesis is that NPAS2-acting within the NAc-contributes to circadian variations in mood, locomotor activity, and motivation. In studies of Drosophila, other groups have shown that mutations in other circadian genes-particularly Clock itself-dramatically alter the fly's responses to drugs of abuse, and we have recently documented clear abnormalities in reward behavior and circadian oscillations in locomotor activity in Clock knockout mice. Interestingly, we have found that Clock is expressed at high levels within the VTA and NAc, suggesting that Clock, like NPAS2, may contribute to circadian variations in the activity of this reward pathway. The goal of the proposed studies is to carry out a systematic evaluation of the role played by NPAS2, Clock, and related circadian gene products expressed in the brain's appetitive circuits in the regulation of mood and motivation. This will be accomplished by investigating mice with mutations in each of these genes. We are also interested in cross talk between these circadian genes and CREB. Thus, the cAMP pathway and CREB are potent regulators of circadian rhythms in Drosophila and in the mammalian SCN, although the mechanisms underlying this regulation are not known, and such regulation has not been explored in other brain regions. Interestingly, we have found dramatic regulation of NPAS2 expression in the NAc by CREB and by stress. We believe that our data establish ideal experimental systems within which we can now investigate the regulation by CREB of circadian oscillations in the brain's reward pathways.